A conserved domain in Escherichia coli Lon protease is involved in substrate discriminator activity

Lon protease of Escherichia coli regulates a diverse set of physiological r esponses including cell division, capsule production, plasmid stability, an d phage replication. Little is known about the mechanism of substrate recog nition by Lon. To examine the interaction of Lon with two of its substrates , RcsA and SulA,,ie generated point mutations in lon which affected its sub strate specificity. The most informative ion mutant overproduced capsular p olysaccharide (RcsA stabilized) yet was resistant to DNA-damaging agents (S ulA degraded). Immunoblots revealed that RcsA protein persisted in this mut ant whereas SuLA protein was rapidly degraded. The mutant contains a single -base change within ion leading to a single amino acid change of glutamate 240 to lysine, E240 is conserved among all Lon isolates and resides in a ch arged domain that has a high probability of adopting a coiled-coil conforma tion, This conformation, implicated in mediating protein-protein interactio ns, appears to confer substrate discriminator activity on Lon, We propose a model suggesting that this coiled-coil domain represents the discriminator site of Lon.